Oxylipins play important roles in stress signaling in plants. The compound 12-oxophytodienoic acid (cis-OPDA) is an early biosynthetic precursor of jasmonic acid (JA), the key phytohormone orchestrating the plant anti-herbivore defense. When consumed by feeding Lepidopteran larvae, plant-derived cis-OPDA suffers rapid isomerization to iso-OPDA in the midgut and is excreted in the frass. Unlike OPDA epimerization (yielding trans-OPDA), the formation of iso-OPDA is enzyme-dependent, and is catalyzed by an inducible glutathione transferase (GSTs) from the larval gut. Purified GST fractions from the gut of Egyptian cotton leafworm (Spodoptera littoralis) and cotton bollworm (Helicoverpa armigera) both exhibited strong OPDA isomerization activity, most likely via transient formation of a glutathione-OPDA conjugate. Out of 16 cytosolic GST proteins cloned from the gut of cotton bollworm larvae and expressed in E. coli, only one catalyzed the OPDA isomerization. The biological function of the double bond shift might be seen in an inactivation of cis-OPDA, similar to the inactivation of prostaglandin A1 to prostaglandin B1 in mammalian tissue. The enzymatic isomerization is particularly widespread among generalist herbivores that have to cope with various amounts of cis-OPDA in their spectrum of host plants.